Abstract
The newly synthesized mutant L501fsX533 Frizzled-4 form and the alpha3beta4 nicotinic acetylcholine receptor expressed in the absence of nicotine accumulate in the endoplasmic reticulum of COS-7 cells and induce the formation of large areas of smooth and highly convoluted cisternae. This results in a generalized block of the transport to the Golgi complex of newly synthesized proteins. Intriguingly, both effects happen peculiarly in COS-7 cells; HeLa, Huh-7, and HEK293 cells expressing the two receptors at similar level than COS-7 cells show normal ER and normal transport toward the plasma membrane. These results question the conclusion that a dominant-negative mechanism would explain the dominance of the mutant L501fsX533 Fz4 allele in the transmission of a form of Familial exudative vitreoretinopathy. Moreover, they indicate that the coordination of endoplasmic reticulum homeostasis in COS-7 cells is particularly error prone. This finding suggests that COS-7 cells may be extremely useful to study the molecular mechanisms regulating endoplasmic reticulum size and architecture.
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Beaumont KA, Shekar SN, Newton RA, James MR, Stow JL, Duffy DL, Sturm RA (2007) Receptor function, dominant negative activity and phenotype correlations for MC1R variant alleles. Hum Mol Genet 16:2249–2260
Borgese N, Francolini M, Snapp E (2006) Endoplasmic reticulum architecture: structures in flux. Curr Opin Cell Biol 18:358–364
Brothers SP, Cornea A, Janovick JA, Conn PM (2004) Human loss-of-function gonadotropin-releasing hormone receptor mutants retain wild-type receptors in the endoplasmic reticulum: molecular basis of the dominant-negative effect. Mol Endocrinol 18:1787–1797
Bulenger S, Marullo S, Bouvier M (2005) Emerging role of homo- and heterodimerization in G-protein-coupled receptor biosynthesis and maturation. Trends Pharmacol Sci 26:131–137
Calebiro D, de Filippis T, Lucchi S, Covino C, Panigone S, Beck-Peccoz P, Dunlap D, Persani L (2005) Intracellular entrapment of wild-type TSH receptor by oligomerization with mutants linked to dominant TSH resistance. Hum Mol Genet 14:2991–3002
D’Agostino M, Tornillo G, Caporaso MG, Barone MV, Ghigo E, Bonatti S, Mottola G (2011) Ligand of Numb proteins LNX1p80 and LNX2 interact with the human glycoprotein CD8alpha and promote its ubiquitylation and endocytosis. J Cell Sci 124:3545–3556
D’Angelo G, Prencipe L, Iodice L, Beznoussenko G, Savarese M, Marra P, Di Tullio G, Martire G, De Matteis MA, Bonatti S (2009) GRASP65 and GRASP55 sequentially promote the transport of C-terminal valine-bearing cargos to and through the Golgi complex. J Biol Chem 284:34849–34860
Frump AL, Lowery JW, Hamid R, Austin ED, de Caestecker M (2013) Abnormal trafficking of endogenously expressed BMPR2 mutant allelic products in patients with heritable pulmonary arterial hypertension. PLoS One 8:e80319
Gariano RF, Gardner TW (2005) Retinal angiogenesis in development and disease. Nature 438:960–966
Guan R, Feng X, Wu X, Zhang M, Zhang X, Hebert TE, Segaloff DL (2009) Bioluminescence resonance energy transfer studies reveal constitutive dimerization of the human lutropin receptor and a lack of correlation between receptor activation and the propensity for dimerization. J Biol Chem 284:7483–7494
Iodice L, Sarnataro S, Bonatti S (2001) The carboxyl-terminal valine is required for transport of glycoprotein CD8 alpha from the endoplasmic reticulum to the intermediate compartment. J Biol Chem 276:28920–28926
Iram SH, Cole SP (2012) Mutation of Glu521 or Glu535 in cytoplasmic loop 5 causes differential misfolding in multiple domains of multidrug and organic anion transporter MRP1 (ABCC1). J Biol Chem 287:7543–7555
Ishii A, Kanaumi T, Sohda M, Misumi Y, Zhang B, Kakinuma N, Haga Y, Watanabe K, Takeda S, Okada M, Ueno S, Kaneko S, Takashima S, Hirose S (2014) Association of nonsense mutation in GABRG2 with abnormal trafficking of GABAA receptors in severe epilepsy. Epilepsy Res 108:420–432
Kaykas A, Yang-Snyder J, Heroux M, Shah KV, Bouvier M, Moon RT (2004) Mutant frizzled 4 associated with vitreoretinopathy traps wild-type frizzled in the endoplasmic reticulum by oligomerization. Nat Cell Biol 6:52–58
Kleizen B, Braakman I (2004) Protein folding and quality control in the endoplasmic reticulum. Curr Opin Cell Biol 16:343–349
Lee SP, Xie Z, Varghese G, Nguyen T, O’Dowd BF, George SR (2000) Oligomerization of dopamine and serotonin receptors. Neuropsychopharmacology 23:S32–S40
Lemma V, D’Agostino M, Caporaso MG, Mallardo M, Oliviero G, Stornaiuolo M, Bonatti S (2013) A disorder-to-order structural transition in the COOH-tail of Fz4 determines misfolding of the L501fsX533-Fz4 mutant. Sci Rep 3:2659
Lotti LV, Torrisi MR, Pascale MC, Bonatti S (1992) Immunocytochemical analysis of the transfer of vesicular stomatitis virus G glycoprotein from the intermediate compartment to the Golgi complex. J Cell Biol 118:43–50
Maiuolo J, Bulotta S, Verderio C, Benfante R, Borgese N (2011) Selective activation of the transcription factor ATF6 mediates endoplasmic reticulum proliferation triggered by a membrane protein. Proc Natl Acad Sci U S A 108:7832–7837
Matsuda S, Yuzaki M (2002) Mutation in hotfoot-4 J mice results in retention of delta2 glutamate receptors in ER. Eur J Neurosci 16:1507–1516
Mazzo F, Pistillo F, Grazioso G, Clementi F, Borgese N, Gotti C, Colombo SF (2013) Nicotine-modulated subunit stoichiometry affects stability and trafficking of alpha3beta4 nicotinic receptor. J Neurosci 33:12316–12328
McElvaine AT, Mayo KE (2006) A dominant-negative human growth hormone-releasing hormone (GHRH) receptor splice variant inhibits GHRH binding. Endocrinology 147:1884–1894
Milhem RM, Ben-Salem SM, Al-Gazali LI, Ali BR (2014) Identification of the cellular mechanisms that modulate trafficking of frizzled family receptor 4 (FZD4) missense mutants associated with FEVR. Invest Ophthalmol Vis Sci 55:3423–3431
Nechamen CA, Dias JA (2003) Point mutations in follitropin receptor result in ER retention. Mol Cell Endocrinol 201:123–131
Ohgane K, Dodo K, Hashimoto Y (2010) Retinobenzaldehydes as proper-trafficking inducers of folding-defective P23H rhodopsin mutant responsible for retinitis pigmentosa. Bioorg Med Chem 18:7022–7028
Orrenius S, Ericsson JL, Ernster L (1965) Phenobarbital-induced synthesis of the microsomal drug-metabolizing enzyme system and its relationship to the proliferation of endoplasmic membranes. A morphological and biochemical study. J Cell Biol 25:627–639
Pascale MC, Erra MC, Malagolini N, Serafini-Cessi F, Leone A, Bonatti S (1992) Post-translational processing of an O-glycosylated protein, the human CD8 glycoprotein, during the intracellular transport to the plasma membrane. J Biol Chem 267:25196–25201
Polishchuk RS, Polishchuk EV, Luini A (2012) Visualizing live dynamics and ultrastructure of intracellular organelles with preembedding correlative light-electron microscopy. Methods Cell Biol 111:21–35
Robitaille J, MacDonald ML, Kaykas A, Sheldahl LC, Zeisler J, Dube MP, Zhang LH, Singaraja RR, Guernsey DL, Zheng B, Siebert LF, Hoskin-Mott A, Trese MT, Pimstone SN, Shastry BS, Moon RT, Hayden MR, Goldberg YP, Samuels ME (2002) Mutant frizzled-4 disrupts retinal angiogenesis in familial exudative vitreoretinopathy. Nat Genet 32:326–330
Sairam MR, Jiang LG, Yarney TA, Khan H (1996) Follitropin signal transduction: alternative splicing of the FSH receptor gene produces a dominant negative form of receptor which inhibits hormone action. Biochem Biophys Res Commun 226:717–722
Saliba RS, Munro PM, Luthert PJ, Cheetham ME (2002) The cellular fate of mutant rhodopsin: quality control, degradation and aggresome formation. J Cell Sci 115:2907–2918
Stornaiuolo M, Lotti LV, Borgese N, Torrisi MR, Mottola G, Martire G, Bonatti S (2003) KDEL and KKXX retrieval signals appended to the same reporter protein determine different trafficking between endoplasmic reticulum, intermediate compartment, and Golgi complex. Mol Biol Cell 14:889–902
Szczesna-Skorupa E, Chen CD, Liu H, Kemper B (2004) Gene expression changes associated with the endoplasmic reticulum stress response induced by microsomal cytochrome p450 overproduction. J Biol Chem 279:13953–13961
Tjeldhorn L, Iversen N, Sandvig K, Bergan J, Sandset PM, Skretting G (2011) Protein C mutation (A267T) results in ER retention and unfolded protein response activation. PLoS One 6:e24009
Ulloa-Aguirre A, Janovick JA, Brothers SP, Conn PM (2004) Pharmacologic rescue of conformationally-defective proteins: implications for the treatment of human disease. Traffic 5:821–837
Vergeres G, Yen TS, Aggeler J, Lausier J, Waskell L (1993) A model system for studying membrane biogenesis. Overexpression of cytochrome b5 in yeast results in marked proliferation of the intracellular membrane. J Cell Sci 106(Pt 1):249–259
Wickert LE, Blanchette JB, Waldschmidt NV, Bertics PJ, Denu JM, Denlinger LC, Lenertz LY (2013) The C-terminus of human nucleotide receptor P2X7 is critical for receptor oligomerization and N-linked glycosylation. PLoS One 8:e63789
Ye X, Wang Y, Nathans J (2010) The norrin/frizzled4 signaling pathway in retinal vascular development and disease. Trends Mol Med 16:417–425
Zhu X, Wess J (1998) Truncated V2 vasopressin receptors as negative regulators of wild-type V2 receptor function. Biochemistry 37:15773–15784
Acknowledgments
This work was supported in part by the Telethon (Grant GGP09029 to SB) and the CNR Research Project on Aging, Regione Lombardia Projects NUTEC ID 30263049 and MbMM-convenzione no. 18099/RCC (SFC and AC). We thank the Monzino Foundation (Milan, Italy) for its generous gift of the Spinning Disk confocal system LCI MLS (PerkinElmer). We thank Cecilia Gotti for the alpha3 and beta4 nicotinic subunit antibodies, Gabriella Caporaso for her continuous support throughout the Project and Nica Borgese for suggestions.
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D’Agostino, M., Crespi, A., Polishchuk, E. et al. ER Reorganization is Remarkably Induced in COS-7 Cells Accumulating Transmembrane Protein Receptors Not Competent for Export from the Endoplasmic Reticulum. J Membrane Biol 247, 1149–1159 (2014). https://doi.org/10.1007/s00232-014-9710-8
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DOI: https://doi.org/10.1007/s00232-014-9710-8